It has been amply documented that the nerve growth factor protein (NGF) has effects on some vertebrate neurons of the peripheral nervous system which are quite indistinguishable from those cellular events which characterize neuronal differentiation. The degree of cellular heterogeniety fo these target neural structures has been a large obstacle in the study of the NGF response. Our laboratory has developed a model system which uses a human clonal diploid cell line called SK-N-SH-SY-5Y (5Y) which exhibits a response to NGF indistinguishable from the NGF response observed in vivo. Using 5Y we plan to carry out an analysis of the NGF response and obtain definitive information as to the degree of correlation existing among those various parameters usually associated with neuronal differentiation. We will also try to reach an understanding as to the nature of NGF binding and internalization in this clone and the correlation between these two events and the cellular events associated with the NGF response. It has been widely suggested that the determination of basal NGF levels in human tissues of interest may have important clinical significance. The presence of biological inhibitors and NGF binding protein in human serum made it necessary for us to develop a more reliable assay as on immunoelectrophoretic techniques. We have also purified human NGF from placenta and plan to complete the characterization of human nerve growth factor (NGF). With antibodies against human beta-NGF, we will determine unequivocally the basal levels of NGF in a number of human tissues of interest. A number of established cell lines have the property that they synthesize and secrete NGF into surrounding media and we have established that sex steroid can induce de novo synthesis and secretion of NGF in the C-6 rat glioma cell line. We plan to develop a better understanding of this mechanism of steroid induction of NGF synthesis. We will also make use of immunoprecipitation techniques so as to isolate the precursor molecule to NGF synthesized in this and other clonal cell lines of interest.